Video rate spectral fluorescence lifetime imaging with a 512×16 SPAD line sensor

Research output: Chapter in Book/Report/Conference proceedingConference contribution


We demonstrate a 512 x 16 CMOS single photon avalanche diode (SPAD) line sensor with per-pixel on-chip histogramming for video rate spectral fluorescence lifetime imaging (sFLIM). On-chip histogramming provides 32-bin histograms per pixel with 11bit/bin dynamic range. In addition, bin widths in time can be programmed from 51.20 ps to 6.55 ns, providing a histogram range from 1.64 ns to 209.72 ns to suit a wide range of fluorescence decays. At the end of a user defined exposure time, the full histogram data (i.e. 32-bins/pixel and 512 pixels) is first transferred to a FPGA in 84.48 μs via 64 data I/O pads at a 33.33 MHz I/O rate. The sensor data is then binned into two user defined spectral bands to provide spectral separation between different fluorophores, before being transferred to a PC via a USB3 connection for further processing. Fluorescence lifetimes for each spectral band are then rapidly estimated in software by applying the Centre-of-Mass Method (CMM), providing two 128 x 128 size spectral lifetime images in 1.384 s (i.e. with a frame rate of 0.72 fps). The frame rate can be increased by reducing the number of bins, reaching a maximum frame rate when only 2 bins are used with the Rapid Lifetime Determination (RLD) algorithm. In this paper we study the lifetime accuracy vs frame rate trade-offs by varying the number of histogram bins while carefully adjusting the bin widths for maximum bin counts. We validate the results using a Rhodamine 110 and Rhodamine B mixture solution which we separate them spectrally by their fluorescence lifetimes.

Original languageEnglish
Title of host publicationHigh-Speed Biomedical Imaging and Spectroscopy IV
EditorsKeisuke Goda, Kevin K. Tsia
ISBN (Electronic)9781510624207
Publication statusPublished - 4 Mar 2019
EventHigh-Speed Biomedical Imaging and Spectroscopy IV 2019 - San Francisco, United States
Duration: 2 Feb 20193 Feb 2019


ConferenceHigh-Speed Biomedical Imaging and Spectroscopy IV 2019
Country/TerritoryUnited States
CitySan Francisco


  • CMM
  • FLIM
  • Fluorescence lifetime
  • On-chip Histogramming
  • RLD
  • Single photon avalanche diode
  • Single Photon Counting
  • SPAD
  • SPC


Dive into the research topics of 'Video rate spectral fluorescence lifetime imaging with a 512×16 SPAD line sensor'. Together they form a unique fingerprint.

Cite this